Cancer Metastasis Rev (2014) 33:217229

DOI 10.1007/s10555-014-9495-3

NON-THEMATIC REVIEW

Calories, carbohydrates, and cancer therapy

with radiation: exploiting the five Rs through
dietary manipulation
Rainer J. Klement & Colin E. Champ

Published online: 17 January 2014

# The Author(s) 2014. This article is published with open access at Springerlink.com

Abstract Aggressive tumors typically demonstrate a high

glycolytic rate, which results in resistance to radiation therapy
and cancer progression via several molecular and physiologic
mechanisms. Intriguingly, many of these mechanisms utilize
the same molecular pathways that are altered through calorie
and/or carbohydrate restriction. Furthermore, poorer prognosis in cancer patients who display a glycolytic phenotype
characterized by metabolic alterations, such as obesity and
diabetes, is now well established, providing another link between metabolic pathways and cancer progression. We review
the possible roles for calorie restriction (CR) and very low
carbohydrate ketogenic diets (KDs) in modulating the five Rs
of radiotherapy to improve the therapeutic window between
tumor control and normal tissue complication probability.
Important mechanisms we discuss include (1) improved
DNA repair in normal, but not tumor cells; (2) inhibition of
tumor cell repopulation through modulation of the PI3KAkt
mTORC1 pathway downstream of insulin and IGF1; (3)
redistribution of normal cells into more radioresistant phases
of the cell cycle; (4) normalization of the tumor vasculature by
targeting hypoxia-inducible factor-1 downstream of the
PI3KAktmTOR pathway; (5) increasing the intrinsic
radioresistance of normal cells through ketone bodies but
decreasing that of tumor cells by targeting glycolysis. These
mechanisms are discussed in the framework of animal and
human studies, taking into account the commonalities and
differences between CR and KDs. We conclude that CR and
R. J. Klement (*)
Department of Radiotherapy and Radiation Oncology, Leopoldina
Hospital Schweinfurt, Gustav-Adolf-Strae 8, 97422 Schweinfurt,
Germany
e-mail: rainer_klement@gmx.de
C. E. Champ
Department of Radiation Oncology, University of Pittsburgh Cancer
Institute, Pittsburgh, PA, USA
e-mail: colinchamp@gmail.com

KDs may act synergistically with radiation therapy for the

treatment of cancer patients and provide some guidelines for
implementing these dietary interventions into clinical practice.
Keywords Calorie restriction . Ketogenic diet . Metabolism .
Radiotherapy

1 Background
Soon after the discovery of X-rays by Wilhelm Conrad
Rntgen in 1895, ionizing radiation was utilized for cancer
treatment. Today, it constitutes the standard of care for many
cancer patients, along with surgery and chemotherapy. Recently, treatment outcomes have been improved in conjunction with a reduction in toxicity through technological innovations such as intensity modulated radiotherapy or stereotactic body radiotherapy. Despite these advancements, several
cancer types continue to elude modern treatment techniques
with radiation therapy (RT). Radioresistance of these tumors
can be ascribed to two factors: environmental and intrinsic.
The former include hypoxia, high lactate levels or the abundance of growth factors within the cellular microenvironment.
Intrinsic factors include chronically activated proliferative,
invasive, and antiapoptotic signaling pathways. A commonality between all of these factors appears to be the upregulation of glycolysis in cancer cells, resulting in the increased
influx of glucose and excessive production of lactate regardless of partial oxygen pressure [13]. This phenomena was
described nearly a century ago [4, 5], known as the Warburg
effect, which affords cells both a high ATP generation and
biomass synthesis [6]. It is the basic principle behind positron
emission tomography (PET) with the glucose analog
2-(18F)fluoro-2-deoxy-D-glucose (FDG). PET studies have
revealed that FDG uptake is inversely correlated with tumor
control probability [7, 8] and overall survival [9], and areas

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with high FDG-PET have been suggested as targets for dose

escalation with dose-painting RT [10].
The Warburg phenotype provides tumors an enhanced
resistance against cytotoxic insults. In fact, work as early as
1933 has revealed that tumor cells have increased ability to
resist radiation damage in the presence of elevated glucose
[11]. However, this may come at the expense of metabolic
flexibility. Hypoxia and genetic defects that chronically drive
proliferation leave such tumors dependent on a steady supply
of nutrients, especially glucose. Additionally, such tumors
appear to benefit from pathological metabolic conditions of
their host, in particular hyperglycemia, hyperinsulinemia, and
elevated insulin-like growth factor (IGF)-1 levels [12, 13]. As
a result, there has been recent enthusiasm towards
metabolism-based therapies targeting whole-body metabolism, cellular kinases and glycolytic enzymes in order to
sensitize these tumors to cytotoxic insults like RT [1416].
As nutrition is a major modulator of global and cellular
metabolism, it becomes apparent that nutritional interventions
may impact cancer progression. In this context, metabolic
targeting via calorie restriction (CR) has been described as a
promising synergistic treatment option [1719]. CR has consistently been shown to extend life span in organisms from
yeast and worms to mice; furthermore, CR protects against
age-related diseases like cancer [20]. While the beneficial
effects of CR on whole-body metabolism, including improved
insulin and glucose profiles, have been described for decades,
recent research has revealed that, on a cellular level, CR
affects the same molecular pathways as current biological
agents proposed for targeting cancer metabolism. Recent data
from our group [21] has revealed that caloric restriction in
mice works synergistically with RT to target and downregulate several of these pathways and to slow tumor growth
(Fig. 1). In humans, as discussed below, these molecular
effects seem to be mediated mainly by the restriction of
carbohydrates (CHOs) rather than total energy, which
provides a rationale for the application of a very low
carbohydrate, high-fat ketogenic diet (KD) in clinical
practice [22]. Yet, the discussion of either CR or the
KD as a low-cost and non-toxic treatment with multiple
molecular targets is lacking in most discussions regarding metabolic targeting strategies.
The goal of this review is to therefore enhance the
awareness for the potential benefits of CR and a KD as
an adjunct to treatment for cancer patients during RT,
and the strong preclinical data revealing that these modalities may enhance the efficacy of RT. Such benefits
range from the cellular level to global metabolism, and
underline the link between tumor cell metabolism and
that of its host. Focus also lies on the commonalities
and differences between these dietary modifications that
should be considered when developing supplemental
dietary treatment strategies.

Cancer Metastasis Rev (2014) 33:217229

Fig. 1 Nutrient deprivation via alternate day fasting (a) or overall caloric
restriction (b) synergistically work with radiation therapy to significantly slow
tumor growth and downregulate several key pathways (c). AL ad libitum
feeding, CR calorie restriction (taken with permission from Saleh et al. [21])

2 Calories or carbohydrates? Similar metabolic effects

of calorie restriction and the ketogenic diet
CR is usually defined as a 3050 % reduction in energy intake
without malnutrition compared to ad libitum feeding. The
caloric deficit can be induced either by intermittent fasting
(IF), the extreme form of which is water-only short term
fasting (STF), or chronic daily energy restriction (DER).
However, as preclinical data is extrapolated to humans for
clinical research design, it is important to point out that DER
in mice corresponds to therapeutic STF in humans. Along

Cancer Metastasis Rev (2014) 33:217229

these lines, fasting for 1 day in the mouse is roughly comparable to a 1-week water-only fast in a human [23].
Protein restriction leading to a negative nitrogen balance
has been shown to mediate the decrease of IGF-1 during CR
[24, 25], explaining the significant decrease in IGF-1 after
STF or the initiation phase of a KD [26], but not after several
weeks of a KD [27] or long-term CR with adequate protein
intake [25]. However, most other metabolic effects of CR
appear to result from the accompanying restriction of CHOs
[28]. KDs were actually developed in the 1920s as a method
of mimicking fasting while avoiding malnourishment in the
treatment of epilepsy [29]. The notion that KDs mimic the
beneficial response to long-term fasting [30, 31] suggests the
possibility to apply this dietary method to the oncological
setting when weight loss must be avoided [22]. As displayed
in Fig. 2, CHO restriction, whether through CR or a KD,
decreases serum glucose and insulin levels, which increases
lipolysis and leads to fatty acid-mediated activation of peroxisome proliferator-activated receptor (PPAR). PPAR
inhibits glycolysis and fatty acid synthesis, while promoting
the transcription of enzymes that increase ketogenesis and
mitochondrial and peroxisomal fatty acid oxidation [32].
The drop in insulin levels that accompanies the reduction in
CHOs lowers the bioavailability of IGF-1 through increased
transcription of IGF binding protein (IGFBP)-1 [33]. When
insulin and free IGF-1 bind to their specific tyrosine kinase
receptors they activate the phosphatidylinositol-3 kinase
(PI3K)Aktmammalian target of rapamycin complex 1
(mTORC1) signaling pathway to promote many of the hallmarks of cancer including sustained proliferative signaling,
resisting cell death and altered cellular metabolism including
increased fermentation of glucose and glutamine [34].
mTORC1 downregulates ketogenesis through its inhibitory
action on PPAR [35]. This action is counteracted during
metabolic stress induced by CR or glucose withdrawal which
decreases the intracellular ATP/AMP ratio and activates liver
kinase B1 (LKB1)adenosine monophosphate-activated protein kinase (AMPK) signaling. AMPK inhibits mTORC1
either directly through phosphorylation of the regulatoryassociated protein of mTOR (Raptor) or indirectly by phosphorylating the mTOR inhibitor tuberous sclerosis complex
protein-2 (TSC2). Increased lipid oxidation resulting from
AMPK activation also increases the NAD+/NADH ratio thus
amplifying the activity of the NAD+-dependent deacetylase
silent mating type information regulation 2 homologue 1
(SIRT1) [36]. SIRT1 influences cellular lifespan and metabolism through epigenetic regulation of gene transcription and
posttranslational protein modifications. Molecular targets of
SIRT1 include LKB1 and peroxisome proliferator-activated
receptor co-activator (PGC1), which is also activated
through AMPK-mediated phosphorylation at Ser538 and
Thr177 and cooperates with PPAR to induce mitochondrial
biogenesis. This was demonstrated recently by Kitada et al.

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[37] in human skeletal muscle cells treated with serum obtained from four healthy obese subjects after a 25 % DER intervention lasting 7 weeks. Compared to treatment with serum
obtained at baseline, there was a significant increase in
AMPK, SIRT1, and PGC1-mediated mitochondrial biogenesis. In addition, significantly higher levels of phosphoAMPK and phospho-SIRT1 were measured in peripheral
blood mononuclear cells compared to baseline. Thus, CR
and CHO withdrawal activate an energy sensing network
consisting of AMPK, SIRT1, PPAR and PGC1 that promotes mitochondrial function and counteracts the insulin/IGF1PI3KAktmTORC1 pathway. Studies by Draznin et al.
[38] and Bergouignan et al. [39] suggest that CHO restriction
alone, and even in the presence of caloric overconsumption, is
sufficient for the activation of this network in human muscle
cells, in line with the finding that AMPK is sensitive not only
to the intracellular ATP/AMP ratio, but also to glycogen stores
[40]. Studies have revealed increased phospho-AMPK levels
in the liver, but not brain of rats fed a KD [41] and in the liver,
but not epidermis or prostate of mice fed a 30 % CR diet [42],
which implies tissue-dependent effects of CHO restriction on
AMPK activation. Nonetheless, Akt and mTOR signaling
were decreased by either the KD or CR in all of these tissue
sites, again indicating the common effects of calorie and CHO
restriction at the cellular level. Thus, CR and likely KDs target
the same molecular pathways that are also targeted individually by drugs to improve cancer treatment outcomes, including
Akt, mTOR, and AMPK (Fig. 2).

3 How calorie and carbohydrate restriction may influence

the response to radiotherapy
Most often, RT is applied in a fractionated fashion with typical
doses per fraction in the range of 1.83 Gy. The biological
rationale behind fractionated RT is based on exploiting the
different responses between fast proliferating tumors and
slowly proliferating normal tissue (Fig. 3). The factors underlying these responses are known as the five Rs of radiobiology (Fig. 4): Repair of sublethal DNA damage; Repopulation
of the tumor; Redistribution of cells to different phases of the
cell cycle; Reoxygenation of hypoxic tumor areas; and finally,
intrinsic Radioresistance as suggested by Steel et al. [43]. The
goal of RT is to utilize these factors in order to maximize the
therapeutic window under the constraints of sufficiently large
tumor control probability (TCP) and acceptable normal tissue
complication probability (NTCP). Any additional intervention
that increases TCP for a given dose while keeping NTCP
constant, decreases NTCP at a given dose while keeping
TCP constant, or both, will likely enhance treatment efficacy
(Fig. 3). However, many pharmaceutical interventions do not
increase the therapeutic window as they are often exceedingly
unspecific and increase both TCP and NTCP at a given

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Cancer Metastasis Rev (2014) 33:217229

Fig. 2 Calorie restriction (CR)

and a ketogenic diet (KD) target
the same molecular pathways that
are also targeted individually by
drugs to improve cancer treatment
outcomes. Arrows indicate
activation, truncated lines
inhibition. Carbohydrate (CHO)
restriction up-regulates fatty acid
oxidation and ketogenesis
(beneficial for normal tissues) and
impairs glycolysis and
glutaminolysis (detrimental to
tumor cells). See Section 2 for
more details

prescribed dose. In contrast, favorable treatment outcomes

through a combination of CR [21, 44] or the KD [45, 46] with
RT have been described in the literature. Data has demonstrated that CR or its pharmaceutical mimetic protects normal cells
and sensitizes cancer cells to various common chemotherapeutic drugs; remarkably, this so-called differential stress resistance was observed across a wide range of normal and
tumor cell lines, mouse strains and even humans [44,
4751]. Apart from their direct relevance for patients undergoing simultaneous chemoradiation, these findings also suggest that CR or the KD may influence the five Rs of radiobiology (Fig. 4) in a manner that increases the therapeutic
window.
3.1 DNA damage repair
The interaction of ionizing radiation with molecules in tissue
leads to the production of free electrons, leaving behind
charged molecules with unpaired valence electrons called
radicals. Radiolysis of water is the most frequent ionization
event outside of the DNA and leads to the formation of
reactive oxygen species (ROS) including the hydroxyl radical
(OH) and its reaction product with oxygen, hydrogen peroxide (H2O2). ROS are able to diffuse to and oxidize DNA at
various sites including the sugar-phosphate backbone leading
to single (SSBs) and double strand breaks (DSBs). While a
single lesion can usually be repaired and is considered sublethal, accumulation of sublethal lesions with increasing dose
can lead to their interaction and conversion to lethal lesions.
Differences between tumors and normal tissues in the ability

to repair sublethal damage are therefore an important rationale

for fractionated RT.
Numerous studies suggest that CR enhances DNA repair of
sublethal damage in normal tissues (reviewed in Ref. [52]),
implying a role for CR in limiting toxicity to normal tissues
during RT. Along these lines, CR may impact DSB repair,
which is vital for cell survival between fractions [53]. The
repair of DSBs is achieved by two different mechanisms
known as non-homologous end joining (NHEJ) and homologous recombination repair (HRR). During NHEJ, the DSB
ends are quickly recognized and bound by the Ku protein
which subsequently recruits the catalytic subunit of the
DNA-dependent protein kinase (DNA-PKcs) to form the
DNA-PK holoenzyme. Binding to DNA triggers the kinase
activity of DNA-PK which recruits and activates other proteins in order to clean and rejoin the DNA ends. Final ligation
is carried out by the interaction of the XRCC4, ligase IV and
XLF proteins. Although it can utilize short homologous sequences of up to 4 bp when possible, NHEJ does not necessarily conserve DNA sequences and is considered error-prone.
In contrast, HRR is an error-free repair mechanism which
requires DNA homology. It is therefore mostly efficient during and shortly after DNA replication in late S and G2 phases
of the cell cycle when sister chromatids are available. It
follows that HRR is an important pathway against DSBinduced lethality in fast proliferating tumors.
A dose of 1 Gy photon irradiation yields approximately
1.000 SSBs and 40 DSBs in a cells nucleus [54], a number
that can be greatly enhanced through the combination with
chemotherapeutic drugs. As noted previously, differential

Cancer Metastasis Rev (2014) 33:217229

221

Fig. 3 Illustration of a typical

tumor control probability (solid
blue line) and normal tissue
complication probability (red
solid line) curve as a function of
total dose delivered to the tumor.
We argue that CR and possibly a
KD may increase the therapeutic
window by favorably affecting
both curves, i.e. a differential
response between tumor and
normal tissue

stress resistance between normal tissue and tumor cells has

been observed when STF was combined with chemotherapy
[44, 4751]. STF likely selectively improves DSB repair in
normal but not cancer cells. In the lung, liver, spleen, and
kidney of aging rats, CR attenuated the decline of NHEJ
activity [55]; this coincided with increased levels of XRCC4
in these tissues. Other NHEJ proteins like XLF and Ku may be
upregulated by CR in a tissue-dependent manner [55, 56].
Like other DNA stress response genes, XRCC4 appears to be
a target of the forkhead box O (FOXO) transcription factor
family [55], which has been implicated with the antitumoral
effects of CR [57]. FOXO-mediated transcription of stress
response proteins is positively regulated by deacetylation

Fig. 4 The five Rs of radiobiology

through SIRT1, while phosphorylation through Akt leads to

its nuclear exclusion and degradation. Furthermore, upon
radiation-induced DNA damage SIRT1 binds to and
deacetylates the repair protein Ku70, which enhances
the efficacy of DSB repair [58]. Thus, CR and possibly
a KD may positively affect NHEJ in normal tissue by
increasing SIRT1 activity and decreasing insulin/IGF-1
PI3KAkt signaling (Fig. 2). These protection mechanisms are likely defective in tumor cells with selfsufficiency in growth signals and constitutively activated
PI3KAkt pathway [50, 59].
Contrary to this, it is possible that CR impairs DSB repair
in tumor cells and thus contributes to increased cell death.
Chen et al. showed that mTOR inhibition through rapamycin
or everolimus impairs both HRR and NHEJ in MCF7 breast
cancer cells, without significant alterations in several important DNA repair proteins [60]. Importantly, a dose-dependent
effect of CR on mTOR inhibition mediated by AMPK was
also observed in a rat model of breast cancer [61], suggesting
that fasting might have similarly negative effects on DNA
repair capacity in mammary tumors as rapamycin. Song et al.
incubated mouse fibrosarcoma cells with 5 mM metformin for
24 h before and after irradiation [62]. The treated cells exhibited a steeper survival curve with a narrower shoulder, indicating increased accumulation of sublethal lesions at a given
dose and suggesting impaired DNA repair.
In summary, CR has been shown to enhance various DNA
repair mechanisms in normal tissues including HRR and
NHEJ, which are essential for RT-induced DSB repair. In
contrast, repair capacity in cancer cells may be left unaffected
or even attenuated through CR. The differential stress resistance between normal and cancerous cells to chemotherapeutic drugs seems to be mediated at least in part by decreased
glucose and free IGF-1 levels [47, 50]; it could therefore be

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speculated that the KD might achieve similar effects, although

this would have to be investigated in future studies.
3.2 Repopulation of the tumor cells
Repopulation, i.e., the cell proliferation occurring during the
course of fractionated RT, occurs in both tumors and normal
tissue, and provides the biological rationale behind altered
fractionation schedules in certain cancer types such as accelerated fractionation in head and neck squamous cell carcinoma or hypofractionation in non-small cell lung cancer [63].
Such cancers respond to RT with an increase in tumor doubling times and hence accelerated proliferation during extended treatment times, therefore decreasing the TCP. Radiobiological modeling suggests that any strategy that delays the
onset and/or decreases the rate of tumor repopulation could
increase TCP at a given NTCP or decrease the late responding
NTCP through the application of smaller doses in the presence
of a larger number of fractions without impairing TCP [64].
Figure 3 demonstrates this effect qualitatively if one assumes
the solid blue curve to be the TCP with accelerated repopulation starting within the treatment period. The dashed blue
curve would indicate the lack of this effect, i.e., a delay in
the onset of accelerated repopulation to a time point after
finishing the treatment. A quantitative calculation for nonsmall cell lung cancer based on the linear-quadratic formalism
was performed by Fowler et al. [65], demonstrating that the
TCP would be roughly doubled for a typical dose of 70 Gy
given in 2-Gy fractions if accelerated repopulation of the
tumor could be delayed long enough.
CR in rodents reduces IGF-1/insulinPI3KAktmTor signaling which has been shown to be correlated with significant
tumor growth delay [21]. CHO restriction in patients with
advanced cancer has also revealed downregulation of this
pathway [66]. The causal and important role of this pathway
in promoting tumor progression is exemplified by the fact that
CR combined with IGF-1 administration [67] or constitutive
PI3K activation through genetic mutations [59] rescues tumors from growth inhibition induced by CR. We recently
reviewed the large number of animal studies showing the
potential of CR [19] and KDs [22] to delay and retard tumor
growth and even metastasis, often without additional treatment. CR in mice is able to slow tumor growth by 5080 %
though it is important to note that the majority of these studies
reduced CHO within the diet and replaced it with fat. It still
remains unclear if and to what extend these findings translate
to humans, the more so as available data suffer from small
sample sizes. A retrospective analysis of five patients with
tuberous sclerosis complex yielded mixed results concerning
tumor progression during a KD and in no case tumor regression was achieved [68]. Other data are more supportive for
targeting tumor cell proliferation through CHO restriction.
Rossi-Fanelli et al. [69] showed that a high-fat diet (80 %

Cancer Metastasis Rev (2014) 33:217229

non-nitrogenous calories from fat) inhibited tumor cell proliferation while a high-dextrose diet (100 % non-nitrogenous
calories from dextrose) increased proliferation over 14 days in
patients with gastrointestinal cancers, though patient numbers
were too small to reach statistical significance. Diets were
administered parenterally and cell proliferation was assessed
using thymidine labeling index on tumor samples, which
measures the fraction of cells in the S phase as a proxy for
de novo DNA synthesis. Zuccoli et al. reported on a female
patient with GBM undergoing two therapeutic fasts followed
by a KD restricted to 600 kcal/day and concomitant RT and
temozolomide treatment [70]. This intervention stopped tumor growth completely as judged by MRI and PET imaging,
but tumor recurrence occurred 10 weeks after suspension of
this diet.
Fast proliferating cancer cells rely on a high glycolytic rate
in order to shuffle phosphometabolites into the pentose phosphate pathway for biosynthesis of nucleic acids and lipids.
Activation of PPAR by KD or CR promotes ketosis and
inhibits glycolysis, therefore abating proliferation in tumor
cells. In normal cells, abundant acetyl-CoA from the breakdown of ketone bodies and fatty acids inhibits glycolysis to
ensure stable ATP levels; tumor cells which often have dysfunction mitochondria lack this flexibility and quickly die
when confronted with glucose withdrawal [7176]. This was
exemplified in a study by Fine and colleagues [77], revealing
that overexpression of uncoupling protein (UCP) 2, a common
defect in tumor mitochondria, rendered these cells vulnerable
to treatment with the ketone body acetoacetate [77]. In these
cells, the decrease in glycolytic ATP production cannot be compensated by oxidative phosphorylation, leading to ATP depletion
and cell growth inhibition. FDG-PET studies in cancer patients
on a KD confirmed that CHO restriction with subsequent insulin
inhibition and ketosis inhibits tumor glycolysis in vivo [66, 70,
78]. The importance of ketone bodies was thereby demonstrated
by Fine and co-workers [66] who found a statistically significant
correlation between the level of ketosis and partial remission or
stable disease on PET scans after a 4-week KD in nine patients
with prior rapid disease progression.
In conclusion, CR and KDs have shown significant
inhibitory effects on tumor growth in animal studies which
would predict a left-shift of the TCP curve (Fig. 3). Based
on mechanistic insights that the IGF-1/insulinPI3KAkt
mTORC1 pathway and glycolysis play a key role for tumor
cell proliferation and supported by positive evidence from
small patient studies we hypothesize that CR and KDs
could be used as supportive strategies to target tumor cell
repopulation during RT.
3.3 Redistribution in the cell cycle
Normal cells interrupt typical cell cycling after exposure to
ionizing radiation in order to allow for enough time for DNA

Cancer Metastasis Rev (2014) 33:217229

repair, or in case of extreme or irreparable damage, prepare for

cell death or senescence. Transition from one phase of the
cycle into the other is regulated by a family of kinases known
as cyclin-dependent kinases (CDKs), whose activity is regulated through three mechanisms: (1) association with phasedependent proteins called cyclins; (2) phosphorylation and dephosphorylation; (3) inhibition by CDK inhibitors such as
p21. Cells are most sensitive to DNA damage during replication and mitosis, i.e., the S and M phases of the cycle, respectively. Therefore, phases preceding mitosis utilize a variety of
molecular pathways known as checkpoints to ensure that
necessary steps for a phase have been completed and no
severe DNA damage has gone unrepaired. In tumor cells,
checkpoints are often overridden by oncogenic activation of
proliferative signaling via PI3K-Akt [79, 80] and/or loss-offunction of gatekeeper genes like TP53. It follows that with
increasing RT fraction number, ionizing radiation leads to a
decreasing fraction of normal cells in sensitive S and M phases
while tumor cells are mostly unaffected by redistribution.
With a mutation rate of more than 50 %, the transcription
factor p53 is the most frequently mutated gene in tumors.
Important transcriptional targets of p53 include p21 and the
growth arrest and DNA-damage-inducible protein Gadd45a,
two CDK inhibitors that promote G1 and G2 arrest, respectively. p53 is strongly connected to the Warburg phenotype
[81] and provides a rationale for the use of cycle-dependent
chemotherapy. p53 mutations disrupt cytochrome c function,
thus decreasing respiration. This leads to compensatory fermentation or the Warburg effect which can be targeted by
glucose restriction. Apontes et al. [82] showed that rapamycin
and metformin acted synergistically to induce G1/G2 arrest
and protect normal cells under both normal and low glucose
conditions against the mitotic inhibitor nocodazole, a drug
causing lethal mitotic arrest; in contrast, the same treatment
did not protect MDA-MB-231 breast cancer cells expressing
mutant p53, and even was toxic under low glucose conditions.
In addition to p53, other frequent mutations in cancer cells are
responsible for constitutive activation of the PI3KAkt pathway. These cells are able to overcome both the G1/S and G2/
M checkpoints normally induced by DNA damage, and continue to divide [79, 80]. However, such cells may selectively
be targeted by glucose withdrawal. Shim et al. [72] showed
that c-Myc transformed cells underwent apoptosis upon glucose restriction, while normal cells remained intact in G0/G1
cell cycle arrest. Glucose restriction was also shown to exert
opposite epigenetic effects upon human telomerase reverse
transcriptase and the cell cycle regulator p16 between immortalized and normal fetal lung fibroblasts, such that the former
underwent apoptosis while the latter responded with an extension of lifespan [74]. The same authors later identified SIRT1
as a key regulator of this mitigation of p16 in normal cells
[83]. Paradoxically, fasting seems to stimulate the translation
of genes involved in growth and proliferation and to further

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increase phosphorylation of Akt in oncogene-activated cells

[51]. However, though this may appear to be adding fuel to
the fire and driving tumor growth, it also demands increased
energy production which eventually leads to an increase in
ROS and cell death under low nutrient and growth factor
conditions [51, 73, 76].
Conversely, in normal cells, the decrease of mitogenic
stimuli induced by CR and perhaps to a lesser extent, the
KD favors redistribution into a non-dividing state in order to
preserve and redistribute energy for cellular protection mechanisms [50]. This finding can be exploited clinically by having
patients fast prior to each RT session and/or chemotherapy
cycle [48, 49]. Safdie et al. reported that fasting before and/or
after chemotherapy decreased symptoms of weakness and
fatigue, while reducing gastrointestinal side effects when compared to a normal diet in six cancer patients undergoing a
median of four cycles of chemotherapy [48]. In C57BL/6J
mice, CR upregulated Gadd45a and p21 in a FOXO1dependent manner [57]. However, tumors with FOXO inactivation due to hyperactive PI3KAkt signaling would be unable to benefit from CR-induced cell cycle arrest under irradiation, providing a further opportunity to widen the therapeutic window.
In summary, CR arranges a redistribution of normal cells in
the cell cycle, potentially protecting them from subsequent
DNA damaging insults like RT. The situation in tumor cells
seems quite contrary. Here, fasting seems to promote cell
cycle progression, M phase accumulation and energy expenditure, in this way rendering such cells synthetically vulnerable to the combination of nutrient restriction with RT or
chemotherapy.
3.4 Reoxygenation
A major challenge for RT is the presence of hypoxic areas
within solid tumors. The lack of oxygen molecules within
these regions inhibits the formation of H2O2 from OH, thus
lessening the frequency and severity of DNA damage. A
single fraction of irradiation preferentially kills the welloxygenated cells, but reoxygenation of hypoxic areas occurs
during fractionated treatment in part due to tumor shrinkage.
Hypoxia facilitates DNA repair and leads to stabilization of
the -subunit of hypoxia-inducible factor (HIF)-1, a transcription factor that lies downstream of mTOR and
upregulates glycolysis [84]. The AktmTOR pathway
upregulates the translation of HIF-1 mRNA in a glucoseand reoxygenation-dependent manner after irradiation [85].
Tumors possess a heterogeneous network of abnormal
blood vessels characterized by chaotic anatomical arrangement, dead ends, and increased leakiness which leads to
increased interstitial fluid pressure [86]. This results in areas
with both chronic and acute hypoxia, the former occurring
where oxygen supply is limited by diffusion from proximal

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Cancer Metastasis Rev (2014) 33:217229

Fig. 5 Proposed workflow of implementing dietary manipulation for cancer patients based on the results from an initial assessment

blood vessels, and the latter where perfusion is transiently

constricted. The abnormal vasculature is caused by an excess
of pro-angiogenic signaling mainly due to vascular endothelial growth factor 2 (VEGF). VEGF is another target of HIF1, but its transcription is also increased through epigenetic
modulation by inflammatory cytokines, growth factors, and
sex hormones. Contrary to what may be expected from
inhibiting VEGF and therefore new blood vessel formation,
evidence has accumulated supporting the hypothesis that antiVEGF therapy actually decreases hypoxia and facilitates the
delivery of chemotherapeutic drugs to cancer cells by normalizing the vasculature which in turn normalizes the microenvironment [86].
Since VEGF is upregulated as a consequence of Akt
mTORHIF-1 signaling, any strategy that inhibits this pathway can be hypothesized to lower VEGF expression and
tumor progression. Mukherjee, Seyfried, and colleagues have
reported that CR downregulates VEGF and normalizes vascularization across a range of several rodent and human prostate and brain tumors [8789]. In the CT-2A mouse astrocytoma, CR increased the perivascular cell coverage of blood
vessels, insinuating decreased leakiness, less interstitial fluid
pressure, and better drug delivery to the tumor [89].
Hyperbaric oxygen therapy (HBOT) is another approach to
overcome hypoxia. The principle of HBOT encompasses
breathing hyperbaric oxygen during irradiation in order to

oxygenate and radiosensitize hypoxic cancer cells. A recent

Cochrane review concluded that HBOT combined with RT
may improve local control in head and neck and cervical
cancers, but at the expense of significant adverse effects
[90]. Recently, Poff et al. evaluated the combination of HBOT
with a KD in the murine VM-M3 model of metastatic cancer
which closely mimics several aggressive human cancers [91].
Interestingly, despite ad libitum feeding, mice on the KD lost
about 10 % body weight, suggesting involuntary under-eating.
While the KD alone increased mean survival time by 57 %,
the combination of HBOT+KD increased survival time by
78 % compared to a standard diet. The translation of these
results into clinical practice remains an open question. It can at
least be hypothesized that ketone bodies might attenuate additional oxidative stress to normal tissues [9294] but not
cancer cells, which are unable to metabolize them [9598].
3.5 Intrinsic radiosensitivity
The Warburg effect seems to be a hallmark of radioresistant
cancer cells. FDG uptake by tumors is a negative predictor of
local control [7, 8] and survival [9], and is employed to guide
the contouring of particularly radioresistant areas for dose
escalation [10]. The high glycolytic rate appears to protect
cancer cells from ROS-induced DNA damage by supplying
large amounts of reducing equivalents like pyruvate, lactate,

Cancer Metastasis Rev (2014) 33:217229

gluthatione, and NAD(P)H that scavenge ROS molecules [1].

Quantifying lactate via bioluminescence imaging in more than
1,000 individual xenografts of human HNSCC, Sattler and
colleagues demonstrated that intra-tumoral lactate concentrations were significantly inversely correlated with tumor control
after a 6-week RT schedule [99]. However, no such correlation
was found for pyruvate, which can be explained by the fact that
its concentration in tumors is much lower than that of lactate.
Ketone bodies and fatty acids inhibit glycolysis [32], which
is why both fasting and the KD have the potential to target the
antioxidative defense mechanisms outlined above. There is
also evidence that due to dysfunctional mitochondrial electron
transport chains, many cancer cells possess high steady state
levels of ROS that quickly lead to cell death once glycolysis is
impaired [46, 73]. On the other hand, oxidation of ketone
bodies in peripheral tissue decreases the NADP+/NADPH
ratio, which increases the amount of reduced gluthatione
available for scavenging H2O2 [93]. This antioxidative property of ketone bodies would not benefit tumor cells which
lack the necessary enzymes to metabolize them [9598].
Furthermore, Shimazu and colleagues showed that betahydroxybutyrate (BHB) levels achievable after a several days
fast or KD potently protected the kidneys of mice from oxidative stress measured by both protein carbonylation and lipid
peroxidation [94]. The action of BHB and, to a lesser extent
acetoacetate, was thereby related to their roles as class I and II
histone deacetylase inhibitors, leading to histone acetylation
with subsequent transcriptional activation of antioxidant
genes like metallothionein and Foxo3a. Finally, the activation of the histone deacetylase SIRT1, which in humans
occurs after CR [37], or more generally, CHO restriction
[38, 39], has been shown to prevent H 2O2 -induced
hyperacetylation of p53 in skeletal muscle cells, therefore protecting against oxidative stress in these tissues
[37].
Cancer stem cells possess the highest intrinsic radiosensitivity and have been implicated in the failure to achieve local
control, yet studies characterizing their metabolic phenotype
are scarce. A recent study by Vlashi et al. suggests that such
cells possess high metabolic flexibility and readily switch
between glycolysis and oxidative phosphorylation if only
one of these pathways is targeted [100]. This might indicate
thatat least in the case of certain gliomasCR or a KD
alone is not sufficient to decrease ATP content and
radioresistance in cancer stem cells.

4 Clinical implementation
Dietary strategies that involve reducing food intake during
cancer treatment leave the treating physician with trepidation
as data has revealed that weight loss during treatment leads to
poorer outcomes [101]. While significant weight loss from

225

CR is a concern, fat loss in overweight patients during and

after treatment may lead to an improved outcome as excessive
adipose tissue in breast cancer patients may help fuel tumor
cells [102]. However, recent data reveals that a CHOrestricted or KD may have a greater effect on attenuating
metabolic factors associated with increased failure rates of
RT, while avoiding the concern of both physician and patient
in regards to severely restricting calories [103].
Most CR studies in animals employ a reduction in calories
by 30 % or greater, and as discussed previously, such a restriction in mice is roughly comparable to a 1 week water-only fast
in humans [23], both options that may not be reasonable for the
cancer patient. This issue may be minimized through IF around
RT treatments, as it results in less weight loss when used for
periods of 23 months [51], similar to RT treatment times.
Other pertinent issues include possible toxicity from CR, as
chronic CR may decrease immune function [104] and impair
wound healing [105], both issues for the post-operative and
immunocompromised patient. Patients on a KD must also be
closely monitored to ensure sufficient vitamins and nutrients
are consumed for immunoprotection and adequate healing.
One of the first CR studies fasted conscientious objectors to
WWII to 1,500 kcal/day while increasing their activity, leading to severe cachexia, malnourishment, and psychological
detriment [106]. Such limits would be similar to those recommendations of 30 % or greater reduction in calories to achieve
CR. While these patients were engaging in activity to increase
their metabolic rate, this may not be dissimilar from the
physiologic state resulting from a metabolically active tumor.
The GBM patient on a KD reported by Zuccoli et al. was
calorically restricted to 600 kcal/day [70]. Such limits on
calories are not feasible in most oncologic settings, and more
reasonable methods to achieve the metabolic effects of CR,
without the potential of severe malnourishment and toxicity,
include IF and CHO restriction [107]. Along these lines,
preclinical data have revealed that the replacement of
CHOs with fat may actually reduce cachexia [108], and
clinical data have shown weight gain in pancreatic [109]
and gastrointestinal [110] cancer patients with fat supplementation. However, patients must be assessed to ensure
they can adequately tolerate a diet exceedingly high in
fat (Fig. 5). We recently found that 5 weeks of a selfprescribed KD in healthy volunteers significantly increased bioelectrical phase angle [111], which is a proxy
for muscle mass and a strong predictor of survival in
cancer patients [112, 113]. Furthermore, randomized dietary studies in noncancer patients have revealed a significant
decrease in blood glucose and the insulin pathway with a noncalorically but CHO-restricted diet versus a low-fat, calorically restricted diet [reviewed in Ref. 114]. Even caloric excess
by 40 % in conjunction with CHO restriction appears to result
in AMPK upregulation, pointing towards CHO and not calories as the prime target of dietary intervention [38].

226

5 Conclusions
Dietary manipulation through CHO restriction, CR, and a KD
may enhance the efficacy of radiation therapy by exploiting
the five Rs of radiotherapy, while simultaneously reducing
treatment-related toxicity. The treating physician, however,
must weigh the benefits and risks of each dietary intervention,
as each may be suitable in varying situations. While there is an
ample amount of preclinical data, and clinical data continues
to accumulate, further studies must take place comparing the
different methods of dietary manipulation during radiation
treatment and assessing their impact on tumor progression.

Conflicts of interest statement No conflicts of interest exist.

Open Access This article is distributed under the terms of the Creative
Commons Attribution License which permits any use, distribution, and
reproduction in any medium, provided the original author(s) and the
source are credited.

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